模拟微重力对人肺微血管内皮细胞凋亡的影响  

Effects of simulated microgravity on the apoptosis of human pulmonary microvascular endothelial cells

作　　者：康春燕[1] 刘长庭[1] 邹琳[1] 袁明[2] 章烨[1] 李天志[1] 王洋[1] 王德龙[1] 刘岩[1] 

机构地区：[1]北京解放军总医院南楼呼吸科,100853 [2]北京中国航天员科研训练中心航天医学基础与应用国家重点实验室,100094

出　　处：《解放军医学杂志》2010年第11期1315-1318,共4页Medical Journal of Chinese People's Liberation Army

基　　金：军队医药卫生“十一五”计划专项滚动课题(06Z048);武器装备预研基金(9140A1704009JB1002);民用航天科研预先研究项目(科工技[2007]302);国家高技术研究发展计划(2008AA12A220)

摘　　要：目的探讨模拟微重力对人肺微血管内皮细胞(HPMEC)凋亡的诱导作用。方法采用回转器模拟微重力环境,将HPMEC分为回转48h组和1g同步对照组。将盛有回转48h组细胞的培养瓶排除气泡后放入细胞回转器,调节转速为30r/min,37℃回转培养;1g同步对照组细胞静置培养相同时间。培养48h后收集两组细胞进行后续检测。在透射电镜下观察细胞超微结构的变化;采用AnnexinV标记的异硫氰酸荧光素(AnnexinV-FITC)对细胞进行染色,在流式细胞仪上检测细胞凋亡率;采用FITC标记的鬼笔环肽(FITC-phalloidin)对纤维状肌动蛋白(F-action)进行标记,在激光共聚焦显微镜下观察细胞内F-actin的变化。结果透射电镜下可见,回转48h组HPMEC呈典型凋亡改变,染色质固缩凝结、边聚于核膜处,细胞器破坏并可见凋亡小体。流式细胞仪检测可见,回转48h组HPMEC的凋亡率(4.04%±0.32%)高于1g同步对照组(1.68%±0.51%,P<0.01)。激光共聚显微镜下可见,1g同步对照组HPMEC内F-actin表达丰富,呈束状,位于胞质内;回转48h组HPMEC的胞质内F-actin的表达明显减少,分布不均,排列紊乱,且细胞核内染色质浓缩凝聚。结论模拟微重力可诱导HPMEC发生凋亡,细胞骨架的破坏可能是模拟微重力诱导其发生凋亡的机制之一。Objective To explore the effects of simulated microgravity on inducing apoptosis of human pulmonary microvascular endothelial cells （HPMECs）. Methods HPMECs were cultured in vitro and divided into 2 groups： 48h clinostat-rotation group （group CR） and control group. The cell culture containers containing HPMECs of group CR were filled with complete medium, and it was fixed onto the clinostat after discharge of bubbles, and then subjected to 30r/min for 48 hours under 37℃. HPMECs of control group were cultured under similar condition but was not subjected to clinostat rotation. Cells of the 2 groups were thereafter collected for the sequential tests. The uhrastructural changes were observed under transmission electron microscope. The HPMECs were stained with AnnexinV labeled fluorescein isothiocyanate （AnnexinV FITC）, and the apoptotic rate was detected by flow cytometry. The fibrous actin （F-actin） was labeled with FITC-phalloidin, and the changes in intracellular F actin were observed under laser scanning confocal microscope （LSCM）. Results Under transmission electron microscope, the typical apoptotic changes were found in group CR HPMECs such as chromatin aggregation peripherally beneath the nuclear membrane, organelle damage and appearance of apoptotic bodies. The results of flow cytometry showed that the apoptotic rate was much higher in group CR （4. 040%±0. 32%） than in control group （1. 68%±0. 51%, P〈0. 01）. It was observed under laser scanning confocal microscopy that the F-actin was expressed abundantly in HPMECs of control group, appearing as thick actin filaments organized into bundles, while the expression of F-actin was obviously decreased in HPMECs, and it was disorganized with chromatin condensation in nuclei. Conclusions Simulated microgravity may induce apoptosis of human pulmonary microvascular endothelial cells. Cytoskeleton damage may be one of the underlying mechanisms of mierogravity induced apoptosis of the endothelial cells.

关 键 词：失重模拟 内皮细胞 毛细血管 细胞凋亡 细胞骨架 

分 类 号：R852.22[医药卫生—航空、航天与航海医学]